Coxiella burnetii, the causative agent of Q fever, poses a significant public health concern worldwide. Diagnosis primarily relies on serological tests. Traditional antigen production methods, typically involving embryonated hen eggs, are labor-intensive, costly, and require biosafety level 3 facilities. In this study, we tested inactivated whole-cell antigens (SAP9 and NMII/AP9) from C. burnetii strains grown in axenic media, offering a safer and more efficient alternative to egg-based production. These antigens were validated using an in-house ELISA method against human patient sera, demonstrating high sensitivity and specificity comparable to ELISA and to the Gold Standard, IFA commercial kits. Notably SAP9 and NMII/AP9 antigens showed no cross-reactivity with intracellular pathogens that cause illness with similar symptoms. This approach represents significant advancement in diagnostic antigen production for Q fever, facilitating cost-effective epidemiological studies and enhancing laboratory safety.

Counting of viable C. burnetii cells by quantitative reverse transcription PCR using a recombinant plasmid (pCB-dotA) as a standard

Background/Objectives: The detection of anti-Coxiella antibodies using serological methods is essential for identifying exposed ruminants and preventing this important zoonotic disease in livestock. In recent years, numerous attempts have been made to increase diagnostic performance as well as simplify the production of serological assays. Commercially available tests often use whole-cell antigens, which can decrease specificity and require high-level biosafety facilities for manufacturing. The aim of this work was to produce three Coxiella burnetii (C. burnetii) antigens in recombinant form and assess them for the detection of anti-Coxiella antibodies in ruminants. Methods: Three recombinant C. burnetii antigens (Com-1, MceB, AdaA) were selected among immunodominant antigens and produced in a heterologous system (Escherichia coli). Following purification, the proteins were utilized to coat ELISA plates and evaluated for seroreactivity against sera from both negative and positive cattle. Results: Com-1 demonstrated the greatest agreement with the commercial test, albeit moderate. MceB exhibited nonspecific reactivity against a large number of sera, while the AdaA showed reactivity against only a few positive sera. Conclusions: Our findings are consistent with previous research, indicating that utilizing a single antigen to identify exposed animals is unfeasible with current knowledge, most likely due to the complex immunological response following C. burnetii infection in cattle. Consequently, it is critical to continue testing and identifying immunoreactive antigens in order to further investigate them and, potentially, select the most appropriate.